High osmolarity improves the electro-transformation efficiency of the gram-positive bacteria Bacillus subtilis and Bacillus licheniformis

A high osmolarity electroporation method has been developed for the efficie nt transformation of Bacillus subtilis and B. licheniformis. The presence o f high concentrations of the osmoticums, sorbitol and mannitol, in the elec troporation, growth and recovery media resulted in an approximately 5000-fo ld increase in the transformation efficiency of B. subtilis, with a maximum value of 1.4 x 10(6) transformants per mu g DNA. The transformation effici ency increased with electric field strength in the range of 12-23 kV/cm usi ng the high osmolarity method. This enhanced transformation efficiency corr elated with improved cell survival rate, indicating that the high osmolarit y media have some protective action on the electrically injured cells. The application of the optimised high osmolarity method to transformation of B. licheniformis resulted in an increase in transformation efficiency by appr oximately 400-folds, with a maximum value of 1.8 x 10(4) transformants per mu g DNA. As with B. subtilis, the highest transformation efficiency of B. licheniformis was achieved at a very high field strength (21 kV/cm). These results indicate that a combination of high osmolarity and high field stren gth may be a useful approach to improving the transformation efficiency of some gram-positive bacterial species which are able to grow in a high osmol arity medium. (C) 1999 Elsevier Science B.V. All rights reserved.